The genus Allochromatium (Chromatiales Chromatiaceae) revisited: a study on its intragenic structure based on multilocus sequence analysis (MLSA) and DNA-DNA hybridization (DDH).

In this study, the taxonomic status of anoxygenic photosynthetic bacteria belonging to the genus Allochromatium is revisited. The inter- and intraspecies relationship of seven Allochromatium strains, including a set of well described type strains, were examined by DNA-DNA hybridization (DDH) and multilocus sequence analysis (MLSA) using segments of seven protein-coding genes. The re-sequencing of the 16S rRNA, the internal transcriber spacer (ITS), multi-gene analysis and DDH comparison indicated that both type strains Allochromatium vinosum DSM 180(T) and Allochromatium minutissimum DSM 1376(T) are closely related to each other forming an independent cluster together with the strains A. vinosum DSM 183 and DSM 1686. The internal comparison of members of this A. vinosum phylogroup showed values of DDH relatedness above 80% and concatenated sequence similarities (4744bp) above 98%. In contrast, the MLSA scheme has identified A. vinosum strain BH-2 as a separate lineage. Strain BH-2 was first classified as a member of the species A. vinosum based on DDH comparison. However, this strain showed the lowest similarity values of the 16S rRNA gene and concatenated sequences, as well as amino acid identity (AAI) when compared to other Allochromatium strains, suggesting that strain BH-2 may represent a new species.

Demequina lutea sp. nov., isolated from a high Arctic permafrost soil.

Two Gram-stain-positive, pigmented, non-motile, non-spore-forming, pleomorphic, rod-shaped bacteria (strains SV45(T) and SV47), isolated from a permafrost soil collected from the Adventdalen valley, Spitsbergen, northern Norway, have been characterized taxonomically using a polyphasic approach. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the two permafrost isolates formed a distinct phyletic line within the suborder Micrococcineae of the order Actinomycetales. DNA-DNA hybridization analyses indicate that strains SV45(T) and SV47 are closely related (60-69 % relatedness) and belong to the same species, although they show slightly different colony pigmentation. The closest phylogenetic neighbour was Demequina aestuarii JC2054(T), with 96 % 16S rRNA gene sequence similarity. Optimum growth of SV45(T) and SV47 occurred aerobically in the absence of NaCl, but both isolates tolerated up to 2 % NaCl (w/v) in the growth medium. Growth under anaerobic conditions was slow and weak. The peptidoglycan of both isolates was of the A4beta type with l-ornithine as the diamino acid and serine as a component of the interpeptide bridge with either d-aspartate (SV45(T)) or d-glutamate (SV47) as the N-terminal amino acid. The major fatty acids present in both isolates were C(15 : 0) (3.2-8.6 %), iso-C(16 : 0) (5.0-8.9 %), anteiso-C(15 : 0) (59.4-61.5 %), anteiso-C(17 : 0) (4.1-8.8 %) and anteiso-C(15 : 1) (4.4-6.4 %). Isoprenoid quinones were present at exceptionally low levels in both isolates, and only demethylmenaquinone DMK-9(H(4)) could be identified with any degree of confidence. Phylogenetic analysis and differences in physiological and biochemical characteristics between the strains and Demequina aestuarii JC2054(T) indicate that these isolates belong to a novel species within the genus Demequina, for which the name Demequina lutea sp. nov. is proposed. The type strain is SV45(T) (=LMG 24795(T) =DSM 19970(T)).

Spirosoma spitsbergense sp. nov. and Spirosoma luteum sp. nov., isolated from a high Arctic permafrost soil, and emended description of the genus Spirosoma.

Two pigmented, Gram-negative, non-motile, pleomorphic rod-shaped bacteria (strains SPM-9(T) and SPM-10(T)) were isolated from a permafrost soil collected from the Adventdalen valley, Spitsbergen, northern Norway. A third isolate (strain M5-H2) was recovered from the same soil sample after the sample had been exposed to simulated Martian environmental conditions. The three strains were characterized taxonomically by using a polyphasic approach. Phylogenetic, chemotaxonomic, physiological and morphological analyses demonstrated that the three isolates were most closely related to members of the genus Spirosoma. 16S rRNA gene sequence data indicated that the three isolates could be divided into two clusters: (i) strain SPM-9(T) and (ii) strains SPM-10(T) and M5-H2. This grouping was confirmed by DNA-DNA hybridization experiments. Strains SPM-9(T) and SPM-10(T) exhibited 92 % 16S rRNA gene sequence similarity to both Spirosoma linguale LMG 10896(T) and Spirosoma rigui WPCB 118(T). The major fatty acids present in all three isolates were summed feature 3 (comprising iso-C(15:0) 2-OH and/or C(16 : 1)omega7c; 43.0-48.2 % of the total), C(16 : 1)omega5c (19.1-21.3 %), C(16 : 0) (6.7-7.3 %), iso-C(17 : 0) 3-OH (4.7-6.0 %) and iso-C(15 : 0) (2.6-5.7 %). On the basis of their phenotypic and genotypic characteristics, the new strains are assigned to two novel species of the genus Spirosoma, for which the names Spirosoma spitsbergense sp. nov. and Spirosoma luteum sp. nov. are proposed. The type strain of Spirosoma spitsbergense is SPM-9(T) (=NCIMB 14407(T)=DSM 19989(T)) and the type strain of Spirosoma luteum is SPM-10(T) (=NCIMB 14406(T)=DSM 19990(T)). An emended description of the genus Spirosoma is also proposed.

Phototrophic purple sulfur bacteria as heat engines in the South Andros Black Hole.

Photosynthetic organisms normally endeavor to optimize the efficiency of their light-harvesting apparatus. However, here we describe two bacterial isolates belonging to the genera Allochromatium and Thiocapsa that demonstrate a novel adaptation by optimizing their external growth conditions at the expense of photosynthetic efficiency. In the South Andros Black Hole, Bahamas, a dense l-m thick layer of these anoxygenic purple sulfur bacteria is present at a depth of 17.8 m. In this layer the water temperature increases sharply to 36 degrees C as a consequence of the low-energy transfer efficiency of their carotenoids (ca. 30%). These include spirilloxanthin, and related polyene molecules and a novel chiral carotenoid identified as spirilloxanthin-2-ol, not previously reported in purple bacteria. To our knowledge, this study presents the first evidence of such a bacterial mass significantly increasing the ambient water temperature. The transduction of light to heat energy to excess heat may provide these anoxygenic phototropic bacteria with a competitive advantage over non-thermotolerant species, which would account for their predominance within the microbial layer.

Viability, diversity and composition of the bacterial community in a high Arctic permafrost soil from Spitsbergen, Northern Norway.

The viable and non-viable fractions of the bacterial community in a 2347-year-old permafrost soil from Spitsbergen were subjected to a comprehensive investigation using culture-independent and culture-dependent methods. LIVE/DEAD BacLight staining revealed that 26% of the total number of bacterial cells were viable. Quantitatively, aerobic microcolonies, aerobic colony-forming units and culturable anaerobic bacteria comprised a minor fraction of the total number of viable bacteria, which underlines the necessity for alternative cultivation approaches in bacterial cryobiology. Sulfate reduction was detected at temperatures between -2 degrees C and 29 degrees C while methanogenesis was not detected. Bacterial diversity was high with 162 operational taxonomic units observed from 800 16S rDNA clone sequences. The 158 pure cultures isolated from the permafrost soil affiliated with 29 different bacterial genera, the majority of which have not previously been isolated from permafrost habitats. Most of the strains isolated were affiliated to the genera Cellulomonas and Arthrobacter and several of the pure cultures were closely related to bacteria reported from other cryohabitats. Characterization of viable bacterial communities in permafrost soils is important as it will enable identification of functionally important groups together with the as yet undescribed adaptations that bacteria have evolved for surviving subzero temperatures for millennia.

Comparison of cultivation-dependent and molecular methods for studying the diversity of anoxygenic purple phototrophs in sediments of an eutrophic brackish lagoon.

Phototrophic anoxygenic purple bacteria play a key role in many aquatic ecosystems by oxidizing sulfur compounds and low-molecular-weight organic compounds using light as energy source. In this study, molecular methods based upon pufM gene (photosynthetic unit forming gene) were compared with culture-dependent methods to investigate anoxygenic purple phototrophic communities in sediments of an eutrophic brackish lagoon. Thirteen strains, belonging to eight different genera of purple phototrophic bacteria were isolated with a large dominance of the metabolically versatile purple non-sulfur bacteria (eight strains), some purple sulfur bacteria (three strains) and two strains belonging to the Roseobacter clade (aerobic phototrophs). The pufM genes amplified from the isolated strains were not detected by the molecular methods [terminal-restriction fragment length polymorphism (T-RFLP)] applied on in situ communities. An environmental clone library of the pufM gene was thus constructed from sediment samples. The results showed that most of the clones probably corresponded to aerobic phototrophic bacteria. Our results demonstrate that the culture-dependent techniques remain the best experimental approach for determining the diversity of phototrophic purple non-sulfur bacteria whereas the molecular approach clearly illustrated the abundance of organisms related to the Roseobacter clade in these eutrophic sediments.

Characterization of purple sulfur bacteria from the South Andros Black Hole cave system: highlights taxonomic problems for ecological studies among the genera Allochromatium and Thiocapsa.

A dense 1 m thick layer of phototrophic purple sulfur bacteria is present at the pycnocline (17.8 m depth) in the meromictic South Andros Black Hole cave system (Bahamas). Two purple sulfur bacteria present in samples collected from this layer have been identified as belonging to the family Chromatiaceae. One isolate (BH-1), pink coloured, is non-motile, non-gas vacuolated, 2-3 microm in diameter and surrounded by a capsule. The other isolates (BH-2 and BH-2.4), reddish-brown coloured, are small celled (4 microm x 2 microm), motile by means of a single polar flagellum. In both isolates (BH-1 and BH-2), the intracellular photosynthetic membranes are of the vesicular type and bacteriochlorophyll a and carotenoids of the normal spirilloxanthin series are present. Both isolates grow well in the presence of sulfide and carbon dioxide in the light. During photoautotrophic growth sulfur globules are stored intracellularly as intermediate oxidation products. According to the 16S rRNA gene sequence data the isolates belong to the genera Thiocapsa and Allochromatium. However, at the species level a number of inconsistencies exist between the phenotypic and phylogenetic data, highlighting taxonomic problems within these genera. These inconsistencies may have implications for microbiologists studying the ecology of anoxygenic phototrophs. For ecologists studying the functioning of an ecosystem it may not be particularly important to know whether a specific isolate belongs to one species or another. However, if one wants to study the role of different populations within a particular functional group then the species concept is important. This study demonstrates that further work is still required on the taxonomy of purple sulfur bacteria in order that microbial ecologists are able to accurately identify a population/species isolated from hitherto undescribed aquatic ecosystems.

Isolation and characterization of spirilloid purple phototrophic bacteria forming red layers in microbial mats of Mediterranean salterns: description of Halorhodospira neutriphila sp. nov. and emendation of the genus Halorhodospira.

Microbial mats developing in the hypersaline lagoons of a commercial saltern in the Salin-de-Giraud (Rhône delta) were found to contain a red layer fully dominated by spirilloid phototrophic purple bacteria underlying a cyanobacterial layer. From this layer four strains of spirilloid purple bacteria were isolated, all of which were extremely halophilic. All strains were isolated by using the same medium under halophilic photolithoheterotrophic conditions. One of them, strain SG 3105 was a purple non-sulfur bacterial strain closely related to Rhodovibrio sodomensis with a 16S rDNA sequence similarity of 98.8%. The three other isolated strains, SG 3301T, SG 3302 and SG 3304, were purple sulfur bacteria and were found to be very similar. The cells were motile by a polar tuft of flagella. Photosynthetic intracytoplasmic membranes of the lamellar stack type contained BChl a and spirilloxanthin as the major carotenoid. Phototrophic growth with sulfide as electron donor was poor; globules of elemental sulfur were present outside the cells. In the presence of sulfide and CO2 good growth occurred with organic substrates. Optimum growth occurred in the presence of 9-12% (w/v) NaCl at neutral pH (optimal pH 6.8-7) and at 30-35 degrees C. The DNA base composition of strains SG 3301T and SG 3304 were 74.5 and 74.1 mol% G + C, respectively. According to the 16S rDNA sequences, strains SG 3301T and SG 3304 belonged to the genus Halorhodospira, but they were sufficiently separated morphologically, physiologically and genetically from other recognized Halorhodospira species to be described as a new species of the genus. They are, therefore, described as Halorhodospira neutriphila sp. nov. with strain SG 3301T as the type strain (=DSM 15116T).

Characterization of three spiral-shaped purple nonsulfur bacteria isolated from coastal lagoon sediments, saline sulfur springs, and microbial mats: emended description of the genus Roseospira and description of Roseospira marina sp. nov., Roseospira navarrensis sp. nov., and Roseospira thiosulfatophila sp. nov.

Three new spirilloid phototrophic purple nonsulfur bacteria were isolated in pure culture from three different environments: strain CE2105 from a brackish lagoon in the Arcachon Bay (Atlantic coast, France), strain SE3104 from a saline sulfur spring in the Pyrenees (Navarra, Spain), and strain AT2115 a microbial mat (Tetiaroa Atoll, Society Islands). Single cells of the three strains were spiral-shaped and highly motile. Their intracellular photosynthetic membranes were of the vesicular type. Bacteriochlorophyll a and carotenoids of the normal spirilloxanthin series were present as photosynthetic pigments. Optimal growth occurred under photoheterotrophic conditions and in the presence of 0.5-4% w/v NaCl. These features are similar to those described for Roseospira mediosalina. Comparative sequence analysis of their 16S rRNA genes placed these strains within the alpha-subclass of Proteobacteria, in a cluster together with Roseospira mediosalina and Rhodospira trueperi. They form a closely related group of slightly to moderately halophilic spiral-shaped purple nonsulfur bacteria.However, the three new isolates exhibited some differences in their physiology and genetic characteristics. Consequently, we propose that they are members of three new species within the genus Roseospira, Roseospira marina sp. nov., Roseospira navarrensis sp. nov., and Roseospira thiosulfatophila sp. nov., with strains CE2105, SE3104, and AT2115 as the type strains, respectively. As a consequence, an emended description of the genus Roseospira is also given.